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Expression of recombinant Thermomonospora fusca xylanase A in Pichia pastoris and xylooligosaccharides released from xylans by it



Expression of recombinant Thermomonospora fusca xylanase A in Pichia pastoris and xylooligosaccharides released from xylans by it



Food chemistry04(3): 1055-1064



The mature peptide of Thermomonospora fusca xylanase A (TfxA) was successfully expressed in Pichia pastoris under the control of AOX1 promoter. The activity of recombinant T. fusca xylanase A (reTfxA) in culture supernatant was 117.3 pl 2.4 U/mg, which is 3 times higher than that of the native TfxA. The optimal temperature and pH for reTfxA were 60 pC and 6.0, respectively. When treated at 70 pC and pH 6.0 for 2 min, the residual activity of the reTfxA was 70%. The reTfxA was very stable over a wide pH range (5.0-9.0). After incubation over pH 5.0-9.0 at 25 pC for 1 h, all the residual activity of reTfxA was over 80%. The Km and kcat values for reTfxA were 2.45 mg/ml and 139 s-1, respectively. HPLC analysis revealed that xylobiose (X2) was the main hydrolysis product released from birchwood xylan and wheat bran insoluble xylan by reTfxA. Hydrolysis results of xylooligosaccharides showed that reTfxA was an endo-acting xylanase and xylobiose, xylotriose (X3), xylotetraose (X4), xylopentaose (X5), and xylohexaose (X6) could be hydrolysed. This is the first report on the expression of reTfxA in yeast and on the determining and quantifying of the hydrolysis products released from xylans and xylooligosaccharides by reTfxA.

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Accession: 013376210

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DOI: 10.1016/j.foodchem.2007.01.028



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